Our goal is to understand how multi-gene clusters are organized and regulated. Specifically, how does the physical organization influence expression of the genes & how is a particular gene within an open cluster chosen for expression? The model evolving from studies of the human beta-globin cluster is that control elements upstream of the genes are essential for locus activation (chromatin opening). These upstream elements also increase the expression of the nearest available genes, with availability chiefly determined by the promoter. This model is supported by the genotype/phenotype correlations observed in patients with deletional beta-thalassemia and hereditary persistence of fetal hemoglobin. We have shown previously that the chicken beta-globin gene, with its 3' enhancer, is expressed in a copy number dependent manner in transgenic mice. The expression level increased approximately 6-fold upon inclusion of the four upstream DNase hypersensitive sites. Each of the four beta- like globins were shown to be under the control of both the upstream hypersensitive sites and the enhancer located between the beta and epsilon genes. To study the effect of the individual upstream hypersensitive sites, we produced lines of mice carrying a single site linked to the beta gene and enhancer. These experiments demonstrated that no single upstream hypersensitive site accounts for the increased beta-globin expression seen in mice containing the complete upstream region. Taken together, our work demonstrates that at least three regions (the beta/epsilon enhancer and at least two upstream sites) plus the promoter are required for proper expression of the individual beta-like globins. Butyrate stimulates the expression of fetal globins in adult cells and is undergoing clinical trials for this purpose. We used transient expression to explore the mechanism of butyrate's actions and found two effects. First, low concentrations of butyrate caused an inhibition of reporter expression. The second effect was stimulation of reporter expression and required higher butyrate concentrations and was promoter-dependent. We postulate that the initial inhibition is due to butyrate-influenced packaging of the transfected DNA into a less transcriptionally active configuration.